Methods for improving seed production in maize

ABSTRACT

Defoliation of maize plants, based on percent seed moisture, has advantageous outcomes including greater number of seeds per pound; increased volume or proportion of saleable seed per field and per female acre; decreased discard of seed due to commercially undesirable size or shape; lower moisture content of seed at harvest; earlier harvest date; less fuel and time expended in drying seed for storage; improved performance in laboratory tests for germination at cold temperatures; improved seed treatment efficacy; improved emergence under stress in field conditions; improved plantability in mechanical systems; more uniform stand; fewer runt plants and improved grain yield.

CROSS-REFERENCE

This utility application claims the benefit of U.S. ProvisionalApplication Ser. No. 61/547,142 filed Oct. 14, 2011, whereinincorporated herein by reference.

FIELD

The discovery is in the field of plant breeding and improvement, relatedspecifically to a production method which provides advantages, forexample, by speeding harvest and increasing the number or proportion ofsaleable seeds produced.

BACKGROUND

Commercial seed production is a process that involves many steps. In thecase of corn (maize), for example, inbred plants must first be crossedin a field under conditions that allow the pollen from the male parentto fertilize the silks on the ear of the female parent in order toproduce the hybrid seeds that will be sold to the farmer for furtherplanting. The quality of this hybrid seed impacts its value in themarketplace.

Many factors can affect the quality of hybrid seed. For example,variations in the moisture content, size, shape, or integrity of theharvested crop at the upstream end of the process may influence theeffectiveness of each further stage of the hybrid seed productionprocess. Substantial variability in the end product may then result andis undesirable. Further, variations in harvested seed at any stage ofproduction may require recalibration of equipment or other operationaladjustments to accommodate the variations. Further still, adjustments toany operating parameter at one step may result in undesirable effectsand/or further adjustments at another step. For example, gravity tablesmay require a relatively consistent flow rate in order to functionproperly, and hence use of surge bins may be needed in order to equalizeflow rates.

Accordingly, there is a need in the art for improved systems and methodsof seed production which result in increased quantity or proportion ofsaleable seed while retaining or improving their germination ability.

The present discoveries relate to these seed production techniques andin particular a technique wherein the plant is treated to inducedefoliation at a particular time in development. In certain embodiments,the timing of treatment is based on the moisture content of thedeveloping seed. When a plant is defoliated at the appropriate stage,production of saleable seed is maximized. Seed that is either too largeor too small is rejected during post-harvest production operations.Therefore, producing more seed of the commercially desired size and/orshape increases productivity per ear and per field. Defoliating a maizeplant at the optimal time based on moisture content of the seed providesimproved control of seed size and/or shape, leading to increasedproductivity in numerous aspects.

Further, seed produced using the disclosed defoliation techniquesexhibits improved performance in stressful growing conditions. Seedsfrom defoliated plants display improved germination and emergence rates,particularly under stress and more uniform stand establishment andhigher grain yield relative to seed produced on non-defoliated plants.

Improved plant productivity is advantageous due to the widespread use ofmaize as human food, livestock feed, and as raw material in industry.The food uses of maize, in addition to human consumption of maizekernels, include both dry- and wet-milling industries. The principalproducts of maize dry milling are grits, meal and flour. The maizewet-milling industry can provide maize starch, maize syrups, anddextrose for food use. Maize oil is recovered from maize germ, which isa by-product of both dry- and wet-milling industries. Maize, includingboth grain and non-grain portions of the plant, is also used extensivelyas livestock feed, primarily for beef cattle, dairy cattle, hogs, andpoultry.

Industrial uses of maize include production of ethanol, maize starch inthe wet-milling industry and maize flour in the dry-milling industry.The industrial applications of maize starch and flour are based onfunctional properties, such as viscosity, film formation, adhesiveproperties and ability to suspend particles. The maize starch and flourhave application in the paper and textile industries. Other industrialuses include applications in adhesives, building materials, foundrybinders, laundry starches, explosives, oil-well muds and other miningapplications.

Plant parts other than the grain of maize are also used in industry: forexample, stalks and husks are made into paper and wallboard and can beused to make cellulosic ethanol. Cobs are used for bedding, for fuel,and to make charcoal.

SUMMARY

Certain embodiments provide a method of improving production in a maizeseed field wherein at least one maize plant is defoliated when themoisture content of the maize plant's developing seed is at or about 35,36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53,54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69 or 70%and the plant is harvested when the moisture content of its developingseed is at or about 10, 11, 12, 13, 14, 15, 16, 17, 18. 19, 20, 21, 22,23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40,41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58,59 or 60%.

Certain embodiments provide a means for determining the optimum time fordefoliation of an inbred. Factors in the determination include, but arenot limited to, environment, genetics and product being utilized.

Use of properly-timed defoliation provides numerous advantages which arediscussed throughout this document, including the Figures and theExamples. These advantages include, and are not limited to, greaternumber of seeds per pound; increased volume or proportion of saleableseed per field and per female acre; decreased discard of seed due tocommercially undesirable size or shape; lower moisture content of seedat harvest; earlier harvest date; less fuel and time expended in dryingseed for storage; improved performance in laboratory tests forgermination at cold temperatures; improved seed treatment efficacy;improved emergence under stress in field conditions; improvedplantability in mechanical systems; more uniform stand and improvedgrain yield.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1—The top bar chart shows that across 472 seed corn fields ofvarious hybrids, the defoliation treatment (“DEFOL” or “DEFOLapplication” or “DEFOL treatment”) resulted in an average of 1821 seedsper pound, which is an increase of 266 kernels, or 17%, over theuntreated check (“DEFOL CHECK”), which yielded an average of 1555 seedsper pound. The difference in kernels per pound between DEFOL anduntreated was significant (P<0.001).

The bottom bar chart shows that across 472 seed corn fields in the U.S.Midwest, the DEFOL treatment increased the proportion of saleable seedby approximately 3.4 percentage points. The graph shows the untreatedsamples had a saleable seed portion of 87.1% whereas the seed harvestedfrom the DEFOL treated area had 90.5% saleable seed. The difference insaleable seed was significant in that P<0.0001.

FIG. 2—The upper left bar chart depicts the difference in percentlarge-seed discard (seed greater than 23/64″ round hole screen) betweentwo treatments from 472 seed fields across the U.S. Midwest. The percentlarge-seed discard decreased from 10.9% to 5.7% with the use of DEFOL,for an overall decrease of large seed discard of 5.2 percentage points.The difference in large seed discard between the two treatments wassignificant in that P<0.0001.

The upper right bar chart depicts the difference in percent small-seeddiscard (seed less than 16.5/64″ round hole screen) between the twotreatments of the seed collected from 472 seed fields across the U.S.Midwest. The percent small seed discard increased from 2.1% in theuntreated check (“DEFOL CHECK”) to 3.9% (“DEFOL”), for an overallincrease of 1.8 percentage points. The difference in small-seed discardpercent was significant in that P<0.0001.

The lower left bar chart depicts total size-based discard, i.e. seed toolarge to sell (large-seed discard) and seed too small to sell(small-seed discard), across the 472 seed fields in the

U.S. Midwest. Total size-based discard decreased from 13% in theuntreated check area to 9.5% in the DEFOL treated area, for an overalldecrease in discard seed of 3.5 percentage points. The difference intotal discard seed between the two treatments was significant in thatP<0.0001.

FIG. 3—This graph depicts the difference in kernels per pound for seedfrom the DEFOL treated area and seed from the untreated area across 472seed fields in the U.S. Midwest. In 95% of the 472 seed fields, thenumber of kernels per pound (“KERLB”) was greater in the DEFOL treatedarea than in the untreated area.

FIG. 4—This graph represents the correlation, and provides a prediction,between the kernels per pound of seed in the untreated area and kernelsper pound of seed in the treated area. The regression equation providesan estimate of the kernels per pound following a DEFOL treatment made atabout 60% developing-seed moisture, based on kernels per pound of theuntreated inbred. For example, if an inbred typically produces seed of1500 kernels per pound, a DEFOL application at around 60% grain moisturewould increase the number of kernels per pound to 1766((1.0382×1500)+208.26).

FIG. 5—This graph depicts the difference in harvest grain moisture inthe seed from the treated area (“DEFOL”) and that of the untreated(“UNT”) area from 313 seed fields in the U.S. Midwest. Bars below the 0x-axis line indicate that the seed in the DEFOL treated area was oflower grain moisture than the seed in its corresponding untreated area.

FIG. 6—This chart depicts the % field emergence (crop establishment) fortreated (“DEFOL”) and untreated (“CHECK”) seeds in stress and non-stressfield locations. Each of the bars represents the average % emergence for16 Pioneer hybrids. Treated and untreated seeds from each hybrid wereproduced in the same seed fields in 2010 (see, Example 1). Seeds wereplanted in the field in spring 2011. Locations were considered stressful(“Stress locs”) if the average emergence of all hybrids was 85% or lessor if the soil temperature remained at 10° C. or below for two weeksafter planting. Locations where average % emergence of all hybrids wasgreater than 85% were considered “Non-stress locs”. The followinglocations were considered “Stress locs”: Eau Claire, Wis.; Janesville,Wis.; Schuyler, Nebr.; Minburn, Iowa; Flandreau, S. Dak. “Non-stress”locs were Johnstown, N. Dak. and Coteau du Lac, Canada.

FIG. 7—This bar chart depicts the % field emergence (crop establishment)for treated (“DEFOL”) and untreated (“CHECK”) seeds from eight Pioneerhybrids. Treated and untreated seeds from each hybrid were produced inthe same seed fields in 2010 (see, Example 1). Seeds were planted inspring 2011 at the following locations: Eau Claire, Wis.; Janesville,Wis.; Schuyler, Nebr.; Minburn, Iowa; Flandreau, S. Dak. The barsrepresent the average % emergence across all locations.

FIG. 8—This bar chart depicts the yield in bushels per acre for treated(“DEFOL”) and untreated (“CHECK”) seeds in two field locations. Each ofthe bars represents the average yield for 15 Pioneer hybrids. Treatedand untreated seeds from each hybrid were produced in the same seedfields in 2010 (see, Example 1). Seeds were planted in spring 2011.

DETAILED DESCRIPTION

The embodiments and aspects discussed herein are described inconjunction with systems, tools and methods which are meant to beexemplary and illustrative, and not limiting in scope. In variousembodiments, one or more of the above-described problems have beenreduced or eliminated, while other embodiments are directed to otherimprovements.

The disclosure of each reference set forth herein is hereby incorporatedby reference in its entirety, to the extent they relate to the materialsand methods described herein. Field crops are bred through techniquesthat take advantage of the plant's method of pollination. In thepractical application of a chosen breeding program, the breeder ofteninitially selects and crosses two or more parental lines, followed byrepeated selfing and selection, thereby producing many unique geneticcombinations. The breeder can theoretically generate billions ofdifferent genetic combinations via crossing, selfing and mutagenesis.Therefore, it is highly unlikely that two breeders will independentlydevelop the same variety having the same traits.

In each cycle of evaluation, the plant breeder selects the germplasm toadvance to the next generation. This germplasm is grown under chosengeographical, climatic and soil conditions and optionally subjected totesting for stress tolerance, insect resistance and other traits.Further selections are then made during and after the growing season.

Production techniques are used to manage the propagation of the maizehybrid, seed for which will be sold into the marketplace. A male inbredand female inbred are generally planted in a field with one or morefemale rows per male row. Plants in the female row are detassled priorto their producing pollen, thus causing the pollen from the male row tofertilize the ears in the female row. Alternatively, the female may bemale sterile and not produce pollen. In any case, the result is thatpollen from the male plant fertilizes the ear on the female plant, thusproducing hybrid seed. This seed is then harvested.

Harvest techniques can have a significant effect on the quality of theseed. For example, harvest must occur at the right time, when the seedis of the appropriate size and developmental stage. Some techniques,such as those disclosed herein, can be used to increase seed quality ator near the time of harvest and/or during post-harvest storage andhandling. These quality attributes may include one or more of thefollowing: greater number of seeds per pound; increased volume orproportion of saleable seed per field and per female acre; decreaseddiscard of seed for commercially undesirable size or shape; lowermoisture content of seed at harvest;

earlier harvest date; less fuel and time expended in drying seed forstorage, which also reduces investment in dryer capacity; improved seedintegrity during post-harvest treatments due to reduced embryo exposurein flatter seeds; improved performance in laboratory tests forgermination at cold temperatures or under other stressful conditions;more uniform coverage of seeds by chemical seed treatments, therebyimproving efficacy of the treatment; reduced weight of theindustry-standard 80,000-kernel bag of seed, providing ergonomicadvantages for an aging farmer population; improved plantability inmechanical systems; improved emergence under stress in field conditions;more uniform stand establishment and improved grain yield.

In one embodiment, the instant disclosure provides optimized treatmentof the female inbred carrying the ear comprising the hybrid seed, inorder to produce a greater number or greater proportion of saleableseed. Saleable seed is that which, among other traits, is viable and isof commercially desirable size and shape.

The defoliation treatment comprises hastening the browning of leafmaterial and stopping or significantly reducing the manufacture ofphotosynthates and/or the movement of photosynthates from the leaf tothe ear. This can be accomplished through, for example, spraying theproduction field with any of a number of salt or herbicide defoliantsolutions, including but not limited to IGNITE®, diquat, paraquat,glyphosate and sodium chlorate. Alternatively or additionally,defoliation may occur by mechanical means, by manual means, or by anycombination of manual, mechanical and/or chemical means. In certainembodiments more than one defoliation treatment may occur. The claimedprocess is not dependent on the method in which the plant is defoliated.Methods are applicable to male-sterile as well as male-fertile plants. Acontrol plant or control field or control treatment will correspond tothe test plant or field except that it does not receive the defoliationtreatment. Previous defoliation methods were designed to improve seedvigor without reducing seed size. See, for example, U.S. Pat. No.6,162,974, at Column 4: “The size of seeds in a maize seed harvest is animportant commercial consideration because larger seed size is preferredin the marketplace.” In contrast, current defoliation methods aredesigned to impact seed size and shape, deliberately producing smaller,flatter, more uniform seed.

Also, previous defoliation methods relied on Growing Degree Days (GDD)or Growing Degree Units (GDU) to determine an appropriate time fordefoliation. Impact of GDD on plant maturation varies across inbreds (asshown in Table 1 of U.S. Pat. No. 6,162,974). In addition, GDD must berecalculated each day of each year and the interaction between GDD andplant maturation is complicated by such factors including but notlimited to soil type and precipitation. Thus a defoliation method basedon GDD is subject to environmental variability.

In contrast, seed moisture level is a stable characteristic, consistentfor an inbred across locations and across years even with varyingweather conditions and therefore is more reliably predictive of theoptimum time for defoliation. Determination of the optimum seed moisturelevel at which to cause defoliation is accomplished through extensivetesting and calibration of the effects of defoliation, encompassingvariables of genetics, treatment type, timing of treatment andenvironmental conditions.

Use of properly-timed defoliation based upon seed moisture contentprovides numerous advantages which include, and are not limited to,greater number of seeds per pound; increased volume or proportion ofsaleable seed per field and per female acre; decreased discard of seeddue to commercially undesirable size or shape; lower moisture content ofseed at harvest; earlier harvest date; less fuel and time expended indrying seed for storage; improved performance in laboratory tests forgermination at cold temperatures; improved seed treatment efficacy;improved emergence under stress in field conditions; improvedplantability in mechanical systems; more uniform stand; fewer runtplants and improved grain yield.

The key factors in that determination (inbred choice) are the startingseed size (kernels per pound) and the stress emergence ability of thefemale (germination). The seed size is a more critical component sincedefoliation can make seeds too small leading to unacceptable levels ofdiscard.

The other component is choosing inbreds that are likely to respond todefoliation by producing hybrids with higher germination and cropestablishment, especially under stressful conditions. Choosing theseinbreds involves characterizing the stress emergence trait for hybridswith these inbreds as females. This is accomplished by evaluating thematerial in a combination of laboratory germination tests andearly-planted field tests. The combined data is used to assign a stressemergence score which characterizes the ability of the material toemerge under stressful conditions. lnbreds that are predicted to producehybrids with below average stress emergence scores are considered asdefoliation candidates.

As used herein and in the appended claims, the singular forms “a”, “an”,and “the” include plural reference unless the context clearly dictatesotherwise. Thus, for example, reference to “a plant” includes aplurality of such plants, reference to “a cell” includes one or morecells, reference to “a seed” includes plurality of such seeds andequivalents thereof known to those skilled in the art, and so forth.

“Determining” or “determination” refers to measuring, assessing,evaluating, estimating, monitoring and/or predicting.

“Defoliant” is any compound that will slow or stop the production ofphotosynthates, and/or movement of photosynthates from the leaf to theear, and includes but is not limited to herbicides and salt compositionsthat may kill the plant. These compositions can also includesurfactants.

“Defoliation” means removal, destruction, or desiccation of asubstantial proportion of leaf tissue of a plant, generally at least50%, up to and including 100%. Defoliation can occur by chemical orphysical (mechanical and/or manual) means, or by any combination of suchmeans. A combination of physical and chemical means of defoliation maybe particularly advantageous in conditions such as drought which mayotherwise result in poor pollination and development of a small numberof large kernels.

The term “emerge” or “emergence” refers to the appearance of a seedlingshoot above the germination medium, for example the soil surface in afield.

The term “emergence rate” refers to a percentage of planted seeds thatemerge. For example, 80% emergence rate indicates 80 of 100 plantedseeds emerge.

“Gene” refers to a nucleic acid fragment that expresses a specificprotein, including regulatory sequences preceding (5′ non-codingsequences) and following (3′ non-coding sequences) the coding sequence.“Native gene” refers to a gene as found in nature with its ownregulatory sequences. “Chimeric gene” refers to any gene that is not anative gene, comprising regulatory and coding sequences that are notfound together in nature. Accordingly, a chimeric gene may compriseregulatory sequences and coding sequences that are derived fromdifferent sources, or regulatory sequences and coding sequences derivedfrom the same source, but arranged in a manner different than that foundin nature. A “foreign” gene refers to a gene not normally found in thehost organism, but that is introduced into the host organism by genetransfer. Foreign genes can comprise native genes inserted into anon-native organism, or chimeric genes. A “transgene” is a gene that hasbeen introduced into the genome by a transformation procedure.

The term “germination” refers to the initial stages in the growth of aseed to form a seedling. A seed is considered germinated if it showssigns of radicle (root) or shoot protrusion or if the emerging seedlingstructures meet specific criteria such as those of the InternationalRules for Seed Testing (International Seed Testing Association, ISTA) orthe Association of Official Seed Analysts, Inc., AOSA).

The term “germinated” may refer to a seed that has produced a viableplant seedling with or without exposure to light in a germinationchamber, growth cabinet, greenhouse or the field.

The term “germination rate” or “germination percentage” refers to apercentage of planted seeds that emerge. For example, 80% emergence rateindicates 80 of 100 planted seeds emerge.

A “hybrid plant” or “hybrid progeny” is an individual produced fromgenetically different parents (i.e., a genetically heterozygous ormostly heterozygous individual). Typically, the parents of a hybriddiffer in several important respects. Hybrids are often more vigorousthan either parent, but they cannot breed true.

The term “hybrid variety” refers to a substantially heterozygous hybridline and minor genetic modifications thereof that retain the overallgenetics of the hybrid line including but not limited to a locusconversion, a mutation or a somoclonal variant.

“Increased germination under stressed conditions” as used herein is ameasure of the seed's ability to germinate under stressed conditions,including but not limited to conditions such as cold, saturated soils,drought, flooding and heat, as compared to a seed of matching geneticsgerminating under the same stressed conditions.

A “plant” can be a whole plant, any part thereof, or a cell or tissueculture derived from a plant. Thus, the term “plant” can refer to anyof: whole plants, plant components or organs (e.g., leaves, stems,roots, etc.), plant tissues, seeds, plant cells and/or progeny of thesame. A plant cell is a cell of a plant, taken from a plant or derivedthrough culture from a cell taken from a plant. Plant cells include,without limitation, cells from seeds, suspension cultures, embryos,meristematic regions, callus tissue, leaves, roots, shoots,gametophytes, sporophytes, pollen and microspores.

The term “progeny” refers to the descendants of a particular plant (selfcross) or pair of plants (cross-pollinated). The descendants can be, forexample, of the F₁, the F₂, or any subsequent generation.

The term “runt(s)” refers to a plant or seedling that is visuallysmaller than the neighboring plants in the field. A plant is considereda runt if it is one or two leaf stages behind the average for thesurrounding plants, or if its height is reduced by one third or morecompared to surrounding plants. The terms “runt(s)” and “uniformity” areoften used to describe similar phenotypes whereby fewer runts areassociated with increased uniformity of seed emergence or standestablishment and vice versa.

“Seed” as used herein refers to a hybrid or inbred plant part comprisingan embryo.

“Stand establishment” as used herein is the ability of a group of seedsto emerge and form normal seedlings, commonly under field conditions.Stand establishment may also be referred to as plant stand or cropestablishment.

“Stress”, “stressed conditions” and related terms refer to any factorsthat decrease plant growth and reproduction below the plant's genotypicpotential including, but are not limited to, cold soil, flooding(submergence), cold rain, frost, snow, soil compaction, and excessiveresidue from previous crops. Stress tolerance refers to the ability of aplant or crop species to withstand stress.

“Seed moisture” can be determined in a number of ways, but often anOhaus© Moisture Balance is employed and used as per manufacturerinstructions. Seed moisture is an important measure as it indicatesmaturation of the seed and affects susceptibility to some diseases.Other methods of determining seed moisture include, but are not limitedto, incubator-drying, drying cabinets, forced air drying, oven drying,microwave drying, sun drying and drying over saturated salt solutions(see, Abendroth, et al., (2011) PMR 1009, Corn Growth and Development,Iowa State University and Winston and Bates, (1960) Ecology 41:232-237.)Various desiccants may be used in combination with seed drying methods.Seeds may be dried at a range of temperatures, including roomtemperature and ambient outdoor temperature, with or without aircirculation.

The present invention further relates to transgenic plant cells andtransgenic plants transformed to contain and express a polynucleotide.“Transformed”, “transfected” or “transgenic” refers to a cell, tissue,organ, or organism into which has been introduced a foreign nucleicacid, such as a recombinant vector. Preferably, the introduced nucleicacid is integrated into the genomic DNA of the recipient cell, tissue,organ or organism such that the introduced nucleic acid is inherited bysubsequent progeny. A “transgenic” or “transformed” cell or organismalso includes progeny of the cell or organism and progeny produced froma breeding program employing such a “transgenic” plant as a parent in across and exhibiting an altered phenotype resulting from the presence ofa recombinant construct or vector. Various methods of planttransformation are currently known and available. For example, theintroduction of DNA sequences into plants and/or plant cells can beaccomplished by Agrobacterium-mediated transformation, viral vectormediated transformation, electroporation and microprojectile bombardmentmediated transformation (particle gun or biolistics methods). The DNAsequence may also be transformed directly into the plastid genome byplastid transformation. As used herein, the term “plastid” means theclass of plant cell organelles that includes amyloplasts, chloroplasts,chromoplasts, elaioplasts, eoplasts, etioplasts, leucoplasts andproplastids. These organelles are self-replicating, and contain what iscommonly referred to as the “chloroplast genome,” a circular DNAmolecule that ranges in size from about 120 to about 217 kb, dependingupon the plant species, and which usually contains an inverted repeatregion.

Transgenic events can include traits for herbicide resistance, drought,yield, oil content of the seed and starch content of the seed, carbonpartitioning within the seed, insecticide resistance or any one or moremyriad other traits.

Seed treatments and coatings may be used on seeds produced using thedisclosed methods. Use of seed coatings and treatments is known in theart; see, for example, U.S. Pat. No. 5,876,739. The combination of thedefoliation treatment and the seed treatment/coating may providedesirable additive effects. Although it is believed that seed treatmentscan be applied to a seed in any physiological state, it is preferredthat the seed be in a sufficiently durable state that no damage isincurred during the treatment process. Typically, the seed would be aseed that had been harvested from the field, removed from the plant andseparated from any other non-seed plant material. The seed wouldpreferably also be biologically stable to the extent that the treatmentwould cause no biological damage to the seed. Better coverage with theseed treatment formulation occurs with the smaller, flatter seedproduced by the methods disclosed herein; in turn, better coverage canprovide improved efficacy. In one embodiment, for example, the treatmentcan be applied to seed corn that has been harvested, cleaned and driedto a moisture content below about 15% by weight. In an alternativeembodiment, the seed can be one that has been dried and then primed withwater and/or another material and then re-dried before or during thetreatment. Within the limitations just described, it is believed thatthe treatment can be applied to the seed at any time between harvest ofthe seed and sowing of the seed.

As used herein, the term “unsown seed” is meant to include seed at anyperiod between the harvest of the seed and the sowing of the seed in theground for the purpose of germination and growth of a plant.

The term “yield” refers to the productivity per unit area of aparticular plant product of commercial value. For example, yield ofmaize is commonly measured in bushels of seed per acre or metric tons ofseed per hectare per season. Yield is affected by both genetic andenvironmental factors. “Agronomics”, “agronomic traits”, and “agronomicperformance” refer to the traits (and underlying genetic elements) of agiven plant variety that contribute to yield over the course of growingseason. Individual agronomic traits include emergence vigor, vegetativevigor, stress tolerance, disease resistance or tolerance, herbicideresistance, branching, flowering, seed set, seed size, seed density,standability, threshability and the like. Yield is, therefore, the finalculmination of all agronomic traits.

Agronomically and commercially important products and/or compositions ofmatter, including but not limited to animal feed, commodities and seedproducts and by-products that are intended for use as food for humanconsumption or for use in compositions that are intended for humanconsumption, including but not limited to flour, meal, syrup, oil,starch, foods containing seeds or seed parts and seed by-products, andthe like are contemplated if these products and compositions of matterare derived from or obtained directly from a seed produced using methodsof the present invention. Such products and/or compositions are alsoreferred to herein as biological samples. The biological samples can bederived from the plant, the plant tissue, or the seed produced by theplant.

All publications and patent applications mentioned in the specificationare indicative of the level of those skilled in the art to which thisinvention pertains. All publications and patent applications are hereinincorporated by reference to the same extent as if each individualpublication or patent application was specifically and individuallyindicated to be incorporated by reference.

Although the foregoing invention has been described in some detail byway of illustration and example for purposes of clarity ofunderstanding, it will be obvious that certain changes and modificationsmay be practiced within the scope of the appended claims.

EXAMPLES

The examples described herein are meant to be representative and asexamples of the discoveries, and are not limiting to the scope of theclaims.

Example 1: Implementation of Defoliation Treatment

In 2010, within 472 seed fields across the Midwest United States, entirefields were treated with Defol® 750 (sodium chlorate from DrexelChemical Company) according to manufacturer's direction and an untreatedcheck strip area was left in each field (i.e., untreated test strip wasnot sprayed with defoliating agent). The check strip area was the widthof a spray boom and the length of the seed field within a representativearea of the seed field. Defol® 750 was applied at 55-60% seed moisturewhich was determined using an Ohaus® MB35 moisture balance. Aftertreatment, seed was allowed to dry naturally in the field untilapproximately at or near physiological maturity (˜32% seed moisture).One to two days prior to machine harvest of the field, all harvestableears from 12 consecutive plants in 3 different areas of each treatment(DEFOL, untreated check) were husked and placed in burlap bags. A tagindicating location, field number, and treatment was placed on theoutside and inside of the bag. A harvestable ear was described as an earequal to or greater than 4 inches in length with more than 30 kernels.

Ears with more than 25% damage from disease or insects were notincluded. Burlap bags were transported to the local production facilityand dried in the lower tunnels of the large production dryers for 3 to 4days (or until approximately 12% seed moisture). Seed moisture at thetime of harvest was determined from the samples collected from eachtreatment area using a DICKEY-john® GAC® moisture tester.

A small plot sheller (AEC small batch sheller) was used to shell seedfrom the cob. Once shelled, the seed was passed over a sample sizerequipped with screens of 23/64 inch round holes (RH) and 16.5/64 inchRH. Seed that remained on top of the 23/64″ RH screen is considered toolarge to sell (large-seed discard). Seed that passes through the 16.5/64RH screen is considered too small to sell and is discarded (small-seeddiscard). Seed that passes through the 23/64″ RH screen but remains ontop of the 16.5/64″ RH screen is considered saleable seed in terms ofsize and shape. From the saleable seed, the number of kernels in 0.25pounds of the saleable seed was counted using a bowl seed counter todetermine kernels per pound. Statistical analysis was performed usingT-test procedures of the Statistical Analysis System (SAS Inst., CaryN.C.). Uniform seed, and particularly flatter, smaller, uniform seed,are preferred for ease and efficiency of treating, packaging, shipping,handling and planting. In addition, seeds that are less round havereduced incidence of “raised-embryo” syndrome. Raised embryos increasesusceptibility to physical damage during seed conditioning and handling.

In addition, defoliation treatment hastens maturation and dry-down sothat harvest can typically occur 4 to 5 days earlier than the untreatedcheck under typical harvest conditions and up to 14 days earlier undercooler weather conditions. This is advantageous in preventing fieldlosses due to inclement weather. It also reduces fuel and dryer capacityrequired for post-harvest drying.

Example 2: Defoliation Treatment Produces Seed with Improved StandEstablishment and Final Grain Yield

Hybrid seeds produced from fields treated with Defol® 750 and fromuntreated check strips were planted in seven field locations in spring2011. Seeds from defoliated and check hybrids were produced in the sameseed fields in 2010. Seeds were planted between Apr. 12 and May 18 inEau Claire, Wis., Janesville, Wis., Schuyler, Nebr., Minburn, Iowa,Flandreau, S. Dak., Johnstown, N. Dak. and Coteau du Lac, Canada. Cropemergence was evaluated at the V3 to V4 stage in all locations andemergence was recorded as the percent of planted seeds that emergenceand produced normal seedlings. For example, if 30 seeds were planted and21 seeds emerged, emergence is reported at 70%.

A location was considered stressful if the average emergence of allhybrids was 85% or less or if the soil temperature remained at 10° C. orbelow for two weeks after planting. Locations where average % emergenceof all hybrids was greater than 85% were considered non-stressful. Thefollowing locations were considered stressful: Eau Claire, Wis.;Janesville, Wis.; Schuyler, Nebr.; Minburn, Iowa; Flandreau, S. Dak.Collectively these locations had average emergence of 74%. Non-stressfullocations were Johnstown, N. Dak. and Coteau du Lac, Canada, whichtogether averaged 91% emergence. On average, seeds from defoliatedhybrids had 10% higher emergence in stressful locations and 4% higheremergence in non-stressful locations, compared to non-defoliated checks(FIG. 6). FIG. 7 shows the difference in % emergence for eightindividual defoliate and check hybrids in the test.

Yield data was collected at the Schuyler, Nebr. and Minburn, Iowa.Fifteen hybrids produced from defoliation and check treatments wereharvested by hand at approximately 18% moisture and the data was pooledacross the two locations, on average, seeds from defoliated hybrids hadapproximately 14 additional bushels per acre at the Schuyler locationand approximately 23 additional bushels per acre at the Minburnlocation.

1-22. (canceled)
 23. A method of improving control of maize seed sizefrom a maize hybrid seed production field, comprising: a. determiningthe percentage of moisture content of a maize hybrid seed from afertilized maize female inbred plant in the hybrid seed productionfield, wherein the field comprises maize female inbred plants that havebeen fertilized by pollen from maize male inbred plants; and b. treatingthe fertilized maize female inbred plants in the hybrid seed productionfield with a means for defoliation, when the percentage seed moisture isbetween 35%-70%, wherein the fertilized maize female inbred plants aredefoliated by said means, thereby improving control of maize seed sizefrom the maize hybrid seed production field.
 24. The method of claim 23,wherein the proportion of flat seed is increased compared to controlseed produced from a field not treated with the means for defoliation.25. The method of claim 23, wherein defoliation is by chemical means.26. The method of claim 23, wherein defoliation is by mechanical means.27. The method of claim 23, wherein multiple defoliation treatments aremade.
 28. The method of claim 23, wherein the seed is harvested from thefield.
 29. The method of claim 28, wherein the seed is harvested atbetween about 20% and 45% moisture.
 30. The method of claim 28, whereinthe seed is removed from the plant and separated from any other non-seedplant material.
 31. The method of claim 23, wherein the percentage seedmoisture is between about 50% and 70% at the time of treating.
 32. Themethod of claim 23, wherein the plant comprises a transgenic eventconferring a trait selected from male sterility, site-specificrecombination, abiotic stress tolerance, altered phosphorus, alteredantioxidants, altered fatty acids, altered essential amino acids,altered carbohydrates, herbicide resistance, insect resistance ordisease resistance.